Disposable cartridge for preparing a sample fluid containing cells for analysis

ABSTRACT

The invention discloses a disposable cartridge for preparing a sample fluid containing cells for analysis. The cartridge comprises one or more parallel preparation units, each preparation unit comprises one or more chambers enclosed between seals and connected in series. Each chamber is configured for receiving an input fluid, performing a procedure affecting the fluid thereby generating an output fluid, and releasing the output fluid. A first chamber of the one or more chambers is a pressable chamber coupled to a first opening, while a last chamber of the one or more chambers is coupled to a second opening. The input fluid of the first chamber is the sample fluid. The one or more preparation units are coupleable to a compartment for performing analysis of the respective output fluids convey able via the second openings.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a national phase application based onPCT/IL2012/000120, filed Mar. 8, 2012, and claims the benefit of U.S.Provisional Application No. 61/450,661, filed Mar. 9, 2011, the contentsof both of which are incorporated herein by reference.

FIELD OF THE INVENTION

The invention relates to the field of performing automatic analysis offluids. More specifically, it relates to a cartridge for preparing asample fluid containing cells for analysis.

BACKGROUND OF THE INVENTION

Point-of-care testing (POCT) is defined as medical testing at or nearthe site of patient care, for example at the doctor's office. Point ofCare Testing systems enable quick performance of tests, for exampleblood tests, eliminating a need for sending samples to laboratory. Quickobtaining of test results allows immediate clinical management decisionsto be made.

It is desirable that such POCT systems be simple to use and requireminimal maintenance. To that end, some systems use fully self-containeddisposable cartridges or strips. In fully-automated systems, nopreliminary sample preparation is required and the cartridges eliminatethe risk of contamination.

U.S. Pat. No. 7,347,617 by Pugia et al., entitled “Mixing inMicrofluidic Devices”, published in 2008, discloses mixing of liquids ina microfluidic device, by dispensing the liquids into a first chamber toproduce combined liquid. The liquids are thereafter discharged throughat least one capillary from the first chamber into a second chamber forcomplete mixing.

U.S. Pat. No. 4,030,888 by Yamamoto et al., entitled “Automatic BloodAnalyzer”, published in 1977, discloses a fully automatic system fordetermining the seven blood parameters. The flow of a diluent and theblood solution starting from the introduction of the sample, to countingportions, means for determination, and to the outlets are controlled bythe supply of either vacuum or pneumatic pressure into two rotaryproportioning cocks and chambers positioned upstream or downstreamthereof.

U.S. Pat. No. 4,826,775 by Burns et al., entitled “Dilution Apparatusand Method”, published in 1989, discloses an automatic dilutionapparatus and method operable in conjunction with automated sampleliquid analysis systems to automatically dilute sample liquids assupplied thereby to automated sample liquid analysis apparatus.

U.S. Pat. No. 4,908,187 by Kipke et al., entitled “Multi-ChamberedPump-Valve Device”, published in 1990, discloses a diluting and mixingdevice which is capable of diluting a first solution to produce a secondsolution which is mixed with an undiluted third solution, toreproducibly produce a unique series of combined solutions. Eachsolution in said series of combined solutions may vary only in theconcentration of a single (selected) reactant, and typically, eachsuccessive solution becomes increasingly more concentrated in theselected reactant. By employing a modification in procedure, eachsuccessive solution in said series may become decreasingly lessconcentrated in the selected reactant. This invention further relates toan automated system comprising the device connected to a stepping motorso as to rapidly and reproducibly produce said series of solutions, saiddevice being further connected to an analyzer means for obtainingchemical, biochemical, or physical chemical data on said series ofsolutions.

U.S. Pat. No. 5,350,693 by Maimon et al., entitled “Multichamber SyringeDevice for Fusing Cells”, published in 1994, discloses an apparatus forfusing cells which includes a multichamber syringe having a firstchamber containing a suspension of cells, a second chamber containing asuspension of cells, and a third chamber containing at least 40% byvolume polyethylene glycol (PEG). The exit passageways of the chambersbeing braided such that the downstream ends thereof are beveled and faceone another at the same level. The relative cross sections of thechambers being of a diameter such that a desired ratio of thesuspensions and solution form in midair a mixture of 15% to 25% PEG byvolume. The apparatus also includes a non-linear tube in fluidcommunication with the syringe for receiving the mixture therefrom and adevice for causing a reciprocating passage of the mixture through thenon-linear tube.

U.S. Pat. No. 5,380,491 by Carver Jr. et al., entitled “Apparatus forPumping and Directing Fluids for Hematology Testing”, published in 1995,discloses an apparatus for hematology testing, which has a sensing unitdefining a counting orifice for the flow of a blood sample through thecounting orifice to analyze the blood sample, and a pump unit havingthree syringes. A first syringe is coupled in fluid communication withthe sensing unit on the inlet side of the counting orifice for injectinga stream of blood sample through the counting orifice. A second syringeis coupled in fluid communication with the sensing chamber on the inletside of the counting orifice for simultaneously injecting a sheath offluid surrounding the sample stream on the inlet side of the countingorifice. A third syringe is coupled to the sensing chamber on the outletside of the counting orifice for aspirating a sheath of fluid from thesensing chamber surrounding the sample stream on the outlet side of thecounting orifice.

U.S. Pat. No. 5,840,254 by Carver Jr. et al., entitled “Apparatus forMixing Fluids for Analysis”, published in 1998, discloses an apparatusfor fluid analysis, such as hematologic analysis, a plurality ofreagent-mixture components are each injected by a respective pumpthrough a valve matrix and into a flow-injection unit. Theflow-injection unit defines a mixing chamber including a plurality ofprotuberances or nubs projecting inwardly toward the center of thechamber, and spaced relative to each other both axially and radially. Asthe reagent-mixture components are injected into the mixing chamber, thenubs agitate the fluid flow and create turbulence, thereby dispersingthe reagent-mixture components and in turn mixing the componentstogether to create a reagent mixture. The flow rates of thereagent-mixture components are adjusted in order to select thereagent-mixture ratio as the components are combined in theflow-injection unit to thereby create the selected reagent mixture. Uponpassage through the flow-injection unit, the reagent mixture is injectedinto a sensing unit for analyzing a particle distribution of themixture.

U.S. Pat. No. 6,241,379 by Larsen et al., entitled “Micromixer Having aMixing Chamber for Mixing two Liquids Through the Use of Laminar Flow,”published in 2001, discloses a micromixer having a mixing chamber formixing two fluids. The mixing chamber has a first inlet arrangement forthe supply of a first fluid and a second inlet arrangement for thesupply of a second fluid. The mixing chamber includes a wall along whichthe first fluid flows, and the second inlet arrangement has at least oneopening in the wall. A projection is located on the wall adjacent to theopening and extending into the mixing chamber so that the first fluidflows around the projection and builds a boundary layer with the secondfluid. Mixing takes place by diffusion through the boundary layer.

U.S. Pat. No. 6,537,813 by Chen et al., entitled “Concurrent Flow MixingMethods and Apparatuses for the Preparation of Gene Therapy Vectors andCompositions Prepared Thereby”, published in 2003, discloses methodsadapted for making mixtures and condensate compositions. In the variousembodiments, it provides controlled and uniform mixing of gene therapyvectors and gene therapy vector vehicles for improved reproducibility,scaleability, stability, and pharmaceutical efficacy.

U.S. Pat. No. 6,820,506 by Kipke et al., entitled “Multi-ChamberedPump-Valve Device”, published in 2004, discloses a multi-chamberedpump-valve device for performing chemical processes, detections oranalyses is described herein. The device includes a plurality ofchambers having variable volumes in fluid communication with one anothervia one or more passageways. Liquid may be directed through the deviceby merely changing the volumes of two or more chambers.

U.S. Pat. No. 6,877,892 by Karp et al., entitled “Multi-StreamMicrofluidic Aperture Mixers”, published in 2005, discloses a robustmicrofluidic mixing devices that mix multiple fluid streams passively,without the use of moving parts. In one embodiment, these devicescontain microfluidic channels that are formed in various layers of athree-dimensional structure. Mixing may be accomplished with variousmanipulations of fluid flow paths and/or contacts between fluid streams.

U.S. Pat. No. 6,915,713 by Kipke et al., entitled “Multi-ChamberedPump-Valve Device”, published in 2005, discloses a multi-chamberedpump-valve device for performing chemical processes, detections oranalyses. The device includes a plurality of chambers having variablevolumes in fluid communication with one another via one or morepassageways. Liquid may be directed through the device by merelychanging the volumes of two or more chambers.

U.S. Pat. No. 6,979,569 by Carver Jr. et al., entitled “Apparatus andMethod for Mixing Fluids for Analysis”, published in 2005, discloses anapparatus for fluid analysis, a plurality of reagent-mixture componentsare each injected by a respective pump through a valve matrix and into aflow-injection unit. The flow-injection unit defines a mixing chamberincluding a plurality of. As the reagent-mixture components are injectedinto the mixing chamber, the nubs agitate the fluid flow, therebydispersing the reagent-mixture components and in turn mixing thecomponents together to create a reagent mixture. The flow rates of thereagent-mixture components are adjusted in order to select thereagent-mixture ratio as the components are combined in theflow-injection unit to thereby create the selected reagent mixture. Uponpassage through the flow-injection unit, the reagent-mixture is injectedinto a sensing unit for analyzing a particle distribution of themixture.

U.S. Pat. No. 7,314,060 by Chen et al., entitled “Fluid Flow ConductingModule”, published in 2008, discloses a fluid flow conducting modulecomprising two or more inlets, one or more outlets, and a chamber thathas a first and second blocks therein. Further, the chamber has agradually wider section in the middle, and two convergent ends. Oneconvergent end is connected to the inlets, and the other convergent endis connected to the outlets. The fluids are injected into the chamberthrough the inlets, flow through the chamber, and conducted towards oneor more outlets for further collection and analysis.

PCT Publication WO/2009/053928 by Shany et al., entitled “Cartridge fora Biological Sample”, published in 2009, discloses a sealed removablecartridge adapted for insertion into n assay device and adapted tocontain a biologic sample, the cartridge comprising two or more assaylocations adapted to facilitate, within said cartridge, two or moreassays of said biologic sample; and an actuator interface adapted tointerface with an actuator of said assay device, to transport saidbiologic sample towards at least one of said assay locations.

U.S. Pat. No. 5,096,669 by Lauks et al., entitled “Disposable sensingdevice for real time fluid analysis”, published in 1992, discloses asystem comprising a disposable device and hand held reader, which canperform a variety of electrochemical measurements on blood or otherfluids. In operation, a fluid sample is drawn into the disposable devicethrough an orifice by capillary action. The orifice is sealed off andthe disposable device is inserted into the reader. The reader whichcontrols the test sequence and flow of fluid causes a calibrant pouchlocated inside the device to be pierced, releasing the calibrant fluidto flow across the sensor arrays to perform calibration. Next an airbladder located in the device is depressed, forcing the sample acrossthe sensors where measurements are performed and read by the readerwhich performs the calibrations. Once the measurements are made, thedevice can be withdrawn from the reader and discarded.

PCT Patent Application WO/2003/044488 by Berndtsson, entitled“Disposable apparatus for use in blood testing”, published in 2003,discloses a disposable apparatus for use in blood testing adapted forsimultaneous dilution of a blood sample into two different dilutionratios. A block-shaped housing has a first and a second receptacle; afirst and a second cylinder, each having a piston moveable therein andeach containing a defined volume of a diluent a valve including a valvebody having three valve body channels extending therethrough and beingpositionable in three distinct positions. In one position thereceptacles are put in simultaneous communication with one each of thecylinders through pairs of the channels. One of the receptacles as afirst means for receiving a blood sample, is adapted to receive a bloodsampling capillary tube.

PCT Patent Application WO/2003/104772 by Larsen, entitled “A disposablecartridge for characterizing particles suspended in a liquid” publishedin 2003, discloses a disposable cartridge for characterizing particlessuspended in a liquid, especially a self-contained disposable cartridgefor single-use analysis, such as for single-use analysis of a smallquantity of whole blood. The self-contained disposable cartridgefacilitates a straightforward testing procedure, which can be performedby most people without any particular education. Furthermore, theapparatus used to perform the test on the cartridge is simple,maintenance free, and portable.

PCT Patent Application WO/2006/084472 by Larsen entitled “Dual samplecartridge and method for characterizing particle in liquid” published in2006, discloses an apparatus for characterizing particles suspended in aliquid, especially a self-contained disposable cartridge for single-useanalysis, such as for single-use analysis of a small quantity of wholeblood. Furthermore, the present invention relates to a method forcharacterizing particles in liquid and a device for sampling a small andaccurate volume of liquid. The apparatus comprises a housing having amixing chamber and a collection chamber separated by a wall containingan opening, a first bore in the outer surface of the housing forentrance of a liquid sample, a first cavity for receiving and holding afirst liquid sample, and a second cavity for receiving and holding asecond liquid sample.

U.S. Pat. No. 6,016,712 by Warden and Kaplan, entitled “Device forreceiving and processing a sample”, published in 2000, discloses adevice for receiving and processing a sample. The device comprises asample receiving element adapted to establish fluid communication withand receive a sample directly from a sample container. The samplereceiving element also allows for introduction of a sample into thedevice. A first chamber is in fluid communication with the samplereceiving element. One or more second chambers are in fluidcommunication with the first chamber. The device also comprises firstand second ports. The first port provides for venting the device. Thesecond port provides for establishing communication between the deviceand means for moving the sample from the sample receiving element to thefirst chamber and for moving the sample from the first chamber to theone or more second chambers. Also included as part of the device ismeans for controlling the precise amount of the sample introduced intoeach of the second chambers. The first chamber and/or one or more of thesecond chambers are adapted for processing the sample. Also disclosedare kits containing the above devices and methods of using the devicesto process a sample.

United States Patent Application 2006/0257993 by Mcdevitt et al.,entitled “Integration of fluids and reagents into self-containedcartridges containing sensor elements”, published in 2006, discloses ananalyte detection device and method related to a portable instrumentsuitable for point-of-care analyses. In some embodiments, a portableinstrument may include a disposable cartridge, an optical detector, asample collection device and/or sample reservoir, reagent deliverysystems, fluid delivery systems, one or more channels, and/or wastereservoirs. Use of a portable instrument may reduce the hazard to anoperator by reducing an operator's contact with a sample for analysis.The device is capable of obtaining diagnostic information usingcellular- and/or particle-based analyses and may be used in conjunctionwith membrane- and/or particle-based analysis cartridges. Analytes,including proteins and cells and/or microbes may be detected using themembrane and/or particle based analysis system.

United States Patent Application 2009/0215072 by Mcdevitt et al.entitled “Methods and compositions related to determination and use ofwhite blood cell counts” published in 2009, discloses an analytedetection device and method related to a portable instrument suitablefor point-of-care analyses. In some embodiments, a portable instrumentmay include a disposable cartridge, an optical detector, a samplecollection device and/or sample reservoir, reagent delivery systems,fluid delivery systems, one or more channels, and/or waste reservoirs.Use of a portable instrument may reduce the hazard to an operator byreducing an operator's contact with a sample for analysis. The device iscapable of obtaining diagnostic information using cellular- and/orparticle-based analyses and may be used in conjunction with membrane-and/or particle-based analysis cartridges. Analytes, including proteinsand cells and/or microbes may be detected using the membrane and/orparticle based analysis system.

PCT Publication WO/2008/149365 by Leshansky et al., entitled “Systemsand Methods for Focusing Particles”, published in 2008, discloses amethod of focusing particles. The method includes: providing asuspension of the particles in a suspending medium; and flowing thesuspension along a channel, such that the flowing suspension occupies acertain volume that has at least one cross-sectional dimension smallerthan 100 μm. The suspending medium has such viscoelastic properties,that flowing the suspension in the channel directs at least some of theparticles towards a focus region, enclosed in said certain volume.

PCT Publication WO/2010/013238, by Bransky et al., entitled“Microfluidic System and Method for Manufacturing the Same”, publishedin 2010, discloses a microfluidic system. The microfluidic systemcomprises a microchannel having in fluid communication with a fluidinlet for receiving a first fluid. The microfluidic system can furthercomprise a piezoelectric actuator which controls the flow of the firstfluid in the microchannel by selectively applying external pressure onthe wall of the microchannel.

SUMMARY OF THE INVENTION

It is one object of the invention to provide a disposable cartridge forpreparing a sample fluid containing cells for analysis, the cartridgecomprising:

-   -   one or more parallel preparation units, each preparation unit        comprising:        -   one or more chambers enclosed between frangible seals and            connected in series, wherein each chamber is configured for            receiving an input fluid, performing a procedure affecting            said fluid thereby generating an output fluid, and releasing            said output fluid;        -   wherein a first chamber of said one or more chambers is a            pressable chamber coupled to a first opening;        -   wherein a last chamber of said one or more chambers is            coupled to a second opening; and    -   wherein the input fluid of said first chamber is said sample        fluid;    -   wherein the one or more preparation units are coupleable to a        compartment for performing analysis of the respective output        fluids conveyable via the second openings.

According to one embodiment the invention provides a disposable whereinsaid one or more pressable chambers included in at least one of said oneor more parallel preparation units include a single chamber being thefirst chamber and the last chamber in said one or more parallelpreparation units.

According to another embodiment the invention provides a disposablewherein said one or more pressable chambers in said one or more parallelpreparation units include each a respective substance.

According to another embodiment the invention provides a disposablecartridge wherein one or more of the chambers of at least one of saidone or more preparation units include interconnected compartments.

According to another embodiment the invention provides a disposablecartridge, wherein the frangible seals enclosing a pressable chamberinclude a preceding frangible seal and a succeeding frangible seal.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the preceding frangible seal of the first chamber prevents flowfrom said first chamber via said first opening.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the succeeding frangible seal of the last chamber prevents flowfrom the respective preparation unit via said second opening.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the first chamber is coupled to the first opening via a firstchannel.

According to another embodiment the invention provides a disposablecartridge, wherein the preceding seal of said first chamber isconfigured for preventing fluid from flowing via a space generatablebetween inner surface of said first channel and a carrier usable tointroduce the sample fluid into said first chamber.

According to another embodiment the invention provides a disposablecartridge, wherein the preceding seal of said first chamber comprises:

-   -   a first frangible seal configured for preventing flow from said        first chamber via said first opening prior to introducing the        sample fluid into said first chamber by the carrier; and    -   a second seal configured for preventing fluid from flowing via        the space subsequent to introducing the sample fluid.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the preceding frangible seal of said first pressable chamber isconfigured to be broken by a carrier usable to introduce the samplefluid into said first chamber.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the preceding frangible seal of said first pressable chamber isconfigured to be broken by the carrier.

According to another embodiment the invention provides a disposablecartridge, wherein the first frangible seal is configured to be brokenby the carrier.

According to another embodiment the invention provides a disposablecartridge, wherein the succeeding frangible seal if configured to bebroken by pressure.

According to another embodiment the invention provides a disposablecartridge, wherein in each one of said one or more parallel preparationunits the last chamber is coupled to the second opening via a secondchannel, the second channel is sealable.

According to another embodiment the invention provides a disposablecartridge, wherein at least one of said one or more parallel preparationunits includes two or more pressable chambers connected in series viaone or more connecting channels, at least one of said one or moreconnecting channels is sealable.

According to another embodiment the invention provides a disposablecartridge, wherein each one of said one or more parallel preparationunits are configured for introducing a carrier usable to introduce thesample fluid directly into a space of said first chamber.

According to another embodiment the invention provides a disposablecartridge, wherein performing a reaction is mixing said input fluid witha substance.

According to another embodiment the invention provides a disposablecartridge, wherein the pressable chambers are configured for mixing byjet flow created by pressure applicable to one or more pressable portionof said pressable chamber.

According to another embodiment the invention provides a disposablecartridge, wherein one or more of the chambers of at least one of saidone or more preparation units include interconnected compartments, atleast one of said interconnected compartments includes a pressableportion.

According to another embodiment the invention provides a disposablecartridge, wherein each compartment includes a pressable portion.

It is an additional object of the invention to provide an analyzingcompartment for obtaining derivatives of a sample fluid and forpresenting said derivatives in a way allowing analysis for the purposeof obtaining parameters of said fluid, the analyzing compartmentcomprising:

-   -   at least one analysis unit, each analysis unit comprising:        -   a hollow member configured for obtaining one or more of said            derivatives and for presenting said derivatives in a way            allowing analysis.

According to one embodiment the invention provides an analyzingcompartment wherein the hollow member of one or more of said at leastone analysis units being configured to obtain a first derivative of saidderivatives for presenting prior to obtaining a second derivative ofsaid derivatives for presenting.

According to another embodiment the invention provides an analyzingcompartment wherein the at least one analysis unit comprises at leasttwo analysis units and wherein said at least two analysis units arecoupled in parallel and configured for presenting a derivative of saidsample fluid in a way allowing to perform in parallel two types ofanalysis.

According to another embodiment the invention provides an analyzingcompartment, wherein the hollow member of one or more of said at leastone analysis unit is a chamber and wherein the one or more of said atleast one analysis unit further comprising:

-   -   a small cross sectioned channel coupled to said chamber, the        small cross sectioned channel is configured for decelerating        flow of a derivative of said sample fluid in the chamber.

According to another embodiment the invention provides a analyzingcompartment, wherein an inner surface of the chamber of the one or moreof said at least one analysis unit has projections coatable with anagent, and wherein the projections are configured to enlarge a contactarea between said agent and said derivative f the sample fluid. It isyet another object of the invention to provide a disposable cartridgefor obtaining a sample fluid containing cells and for preparing thesample fluid for analysis, the cartridge comprising:

-   -   a series of at least two connected chambers, amongst said at        least two chambers are a first chamber and a last chamber, each        chamber in said series is enclosed between frangible seals and        is configured to perform a procedure whose input is a first        derivative of the sample fluid and whose output is a second        derivative of said sample fluid;    -   wherein the chambers in said series are configured for        performing consecutive procedures,    -   wherein the first derivative obtainable by the first chamber is        the sample fluid; the first derivative obtainable by each one of        all chambers apart of said first chamber is the second        derivative respective of the preceding chamber in the series.

According to one embodiment the invention provides a disposablecartridge, wherein the seals comprising:

-   -   a preceding seal configured for sealing a respective chamber        prior to obtaining the first derivative; and    -   a succeeding seal configured to be breached for releasing the        second derivative from the respective chamber.

According to another embodiment the invention provides a disposablecartridge, wherein the preceding seal is a frangible seal.

According to another embodiment the invention provides a disposablecartridge, wherein the preceding seal is a re-sealable seal.

According to another embodiment the invention provides a disposablecartridge, wherein the succeeding seal is a frangible seal.

BRIEF DESCRIPTION OF THE DRAWINGS

In order to understand the invention and to see how it may be carriedout in practice, embodiments will now be described, by way ofnon-limiting example only, with reference to the accompanying drawings,in which:

FIG. 1 schematically illustrates a system for analysis of a sample fluidusing the cartridge, according to certain embodiments of the invention;

FIG. 2 schematically illustrates a cartridge already containing the bodyfluid as inserted into cartridge holding unit, according to certainembodiments of the invention;

FIG. 3 provides a detailed illustration of a cartridge, according to acertain embodiments of the invention;

FIGS. 4A and 4B depict a preceding seal, according to certainembodiments of the invention;

FIGS. 5A and 5B depict preceding seal alternative to the sealillustrated in FIGS. 4A and 4B, according to certain embodiments of theinvention;

FIGS. 6A and 6B depict another alternative preceding seal, according tocertain embodiments of the invention;

FIG. 7 presents a cartridge comprising a chamber containing twocompartments, according to certain embodiments of the invention.

FIG. 8 presents a cartridge comprising a preparation unit composed oftwo chambers, according to certain embodiments of the invention;

FIGS. 9A and 9B presents two configurations of a cartridge comprisingmore than one preparation unit, according to certain embodiments of theinvention;

FIG. 10 schematically illustrates an analyzing compartment, according tocertain embodiments of the invention;

FIG. 11 schematically illustrates an alternative analyzing compartment,according to certain embodiments of the invention;

FIG. 12 schematically illustrates an analyzing compartment, comprisingtwo analyzing units, according to certain embodiments of the invention;

FIGS. 13A and 13B schematically illustrate a cartridge comprising apreparation compartment and an analyzing compartment, according todifferent embodiments of the invention; and

FIGS. 14A, 14B and 14C schematically depict samplers according tocertain embodiments of the invention.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

In the following description components that are common to more than onefigure will be referenced by the same reference numerals.

In addition, unless specifically noted, embodiments described orreferenced in the present description can be additional and/oralternative to any other embodiment described or referenced therein.

The object of this invention is to provide a cartridge for preparing asample fluid containing cells for analysis. The sample fluid may be abody fluid, for example: blood, cerebrospinal fluid (CSF), pericardialfluid, pleural fluid, or any other fluid that may contain cells. Cellsmay be any type of prokaryotic cells, for example bacteria; eukaryoticcells, for example red blood cells; white blood cells (Leukocytes);epithelial cells; circulating tumor cells; cellular fragments, forexample platelets; or others.

For the purpose of explaining the invention and due to consideration ofsimplicity, a cartridge for preparing blood sample for optical analysisresulting in obtaining a Complete Blood Count (CBC) is referencedthroughout the description of the present invention. However, it shouldbe appreciated that the invention is not limited to CBC. Disposablecartridges in accordance with the invention may be used for multipleapplications where analysis of cells is desired, such as HIV monitoring(such as using CD4/CD8 ratio), detection of f-hemoglobin, Malariaantigen or other blood parasites, Paroxysmal Nocturnal Hemoglobinuria(PNH), diagnosis of Celiac disease using Intestinal EndomysialAutoantibodies (EmA), Alzheimer's disease, or any other application forwhich cell-based diagnosis is relevant.

FIG. 1 schematically illustrates a system 101 for analysis of a samplefluid using a cartridge 102, according to certain embodiments of theinvention. For example, the system 101 may be usable as a Point of CareTesting (POCT) system which enables quick obtaining of laboratoryresults in a doctor's office. The system 101 comprises a cartridgeholding unit 103, a pump 104, and an analyzing module 105 comprising adata processing unit 106. The analyzing module 105 may be configured toperform an analysis, e.g., optical analysis and/or electrical impedanceanalysis etc. Accordingly, the module may comprise a suitable sensingelement 107 configured for detecting and measuring parameters used foranalysis. For example, optical sensor (such as a CCD, CMOS orphoto-multiplier) can be used in an analysis module configured foroptical analysis. The module may also comprise an excitation member 108,such as a light source for emitting light of a pre-determined wavelength suitable for the required type of analysis of the sample fluid.The excitation member 108 is possibly coupled to the sensor 107, e.g.,in order to synchronize operations thereof. Also coupled to the sensor107 is the data processing unit 106, that serves for processing andstoring data acquired by a analysis module. The pump 104 serves forgenerating a pressure gradient, such as vacuum, that drives a flow of asample fluid inside the cartridge.

In certain embodiments of the invention the system is configured forperforming a complete blood count. In these embodiments the sensor 107may be a camera which takes images of cells flowing inside the cartridge(as explained below with reference to the following figures). Acquiredimages are then processed by the data processing unit using suitablesoftware and/or hardware in order to determine number of cellscorresponding to each blood cell type (e.g., neutrophils, lymphocytes,erythrocytes, etc.) present in an analyzed blood sample.

FIG. 2 schematically illustrates a cartridge 201 according to certainembodiments of the invention. A sampler 202, serving for theintroduction of a sample fluid into the cartridge is inserted into thecartridge 201 from one side. An analyzing compartment 203 is coupled tothe cartridge 201 from the other side. While the preparation of thesample fluid for analysis is performed by the cartridge, presenting ofthe prepared sample fluid in a way allowing its analysis by the system101 is performed by the analyzing compartment.

In a described embodiment the cartridge and the analyzing compartmentare coupled. The cartridge and the analyzing compartment may bemanufactured together and coupled during, or immediately aftermanufacturing, or they may be manufactured separately and become coupledprior to marketing the cartridge to its end user or even just prior tousage thereof, possibly even by a person performing the test orautomatically inside system 101.

Although in FIG. 2 the cartridge and the analyzing compartment appear tobe two separate compartments coupled together, this is non-limiting, andin other embodiments the analyzing compartment may comprise anintegrative part of a cartridge. Furthermore, while in the presentembodiment the analyzing compartment 203 appears as coupled to thecartridge, i.e., it is not an part thereof, in certain embodiments theanalyzing compartment 203 may be considered as part of the cartridgewhile 201 relates to a “preparation compartment”. That is, a cartridge204 according to such embodiments may be composed of a preparationcompartment 201 and an analyzing compartment 203. The cartridge 204 mayalso, according to certain embodiments, comprise a preparationcompartment 201, and be coupleable to an analysis compartment.Hereinafter, due to simplicity considerations, the description refers tocartridge 201. However, it should be considered that although notspecifically written, reference is made, mutatis mutandis, to cartridge204 as well.

While in the embodiment illustrated in FIG. 2 the sampler 202 and theanalyzing compartment 203 appear to be on both sides of the cartridge,this is non-limiting as well. According to other embodiments the samplerand the analyzing compartment may be positioned, with reference to thecartridge 201, in any way applicable to the case. For example, theanalyzing compartment 203 may be positioned above or below the cartridge201, on its side, on the side where the sampler 202 is positioned, oreven in a gap, or a window, inside the cartridge.

Relating to the sampler 202, it is described below, with reference toFIG. 14. Yet, it should be mentioned here that the sampler is not adirect part of the cartridge 201 (or 204). It is a separate memberhaving a carrier for holding the sample fluid, while the carrier can be,for example, a capillary. According to certain embodiments, the system101 automatically couples the sampler 201 to the cartridge 201 or 204 inorder to introduce the sample fluid thereto.

According to certain embodiments, the sampler may be considered as partof the cartridge, e.g., by coupling the sampler to the cartridge usingany available mean such as a coupling-strip. Such a coupled sampler maybe detached from the carrier (the capillary) in order to prevent it frombreaking. Alternatively, it is possible to consider the sampler as partof the cartridge further to insertion of the carrier thereto.

FIG. 3 provides a detailed illustration of a cartridge, according tocertain embodiments of the invention. In the cartridge 201, a firstopening 301, located in one of the sides thereof, is configured forreceiving the carrier carrying a sample fluid. A first channel 302 iscoupled to the first opening 301 and to a chamber 303. The chamber 303is configured to receive the sample fluid and to perform a procedureaffecting it, thereby forming an output fluid. Then, the chamber isconfigured to release the output fluid into the second channel 304, andtherefrom out of the cartridge via a second opening 305. A precedingseal 306, configured to prevent flow from the chamber via the firstopening is coupled to the first channel 302 while a succeeding seal 307,configured to prevent flow from the chamber via the second opening iscoupled to the second channel 304.

It was previously explained that the term “output fluid” is used forreferring to the fluid being the result of the procedure affecting thesample fluid, wherein the output fluid is further conveyed from thechamber. Hence, analogously, the fluid entering the chamber, prior toaffecting the procedure, is referred to as “input fluid”. It can be thenobserved that the input fluid of a chamber is the sample fluidintroduced thereto.

In FIG. 3 the first and second openings are illustrated when they arepositioned opposite one to the other. It should be appreciated thoughthat this is non-limiting as the two openings may be perpendicularlypositioned, for example. Any other relative position of the opening isallowed, with the necessary modification, including also positioning thetwo openings in the same side of the cartridge.

The procedure affecting a sample fluid, performed inside a chamber, suchas chamber 303, may be any procedure resulting with a change of aphysical or a chemical state of the sample fluid or of the cellscontained within the sample fluid. Examples of possible procedures areheating, mixing, diluting, staining, permeabelization, lysis, etc. Someof the procedures will be described below with reference to thefollowing figures.

In certain embodiments of the invention the chamber 303 is pre-loadedwith a substance. The pre-loaded substance may be a liquid substance, asolid substance or a combination thereof. The substance may consist of asingle reagent or of several different reagents. An example of a liquidsubstance consisting of several reagents is PBS (Phosphate BufferedSaline), while examples of solid substances are lyophilized antibodies,different kind of powdered stains dissolvable, e.g., in water or inethanol, coated beads, etc. A substance may be lying free on the bottomof the chamber or may be attached to the inner surface of the chamber.Alternatively, a substance may be attached to a filling, such as spongeor microfibers, filling the space of the chamber, enlarging the surfacearea exposed to the sample fluid.

Furthermore, some possible procedures, such as heating, do not requirehaving a pre-loaded substance in the chamber. Therefore, in certainembodiments the chamber is not pre-loaded with a substance, while it ispossible that the chamber holds instead (or in addition to a pre-loadedsubstance) some sort of mechanism, such as a heating mechanism or partthereof. In addition, understanding that pre-loading the substance maybe performed while manufacturing the cartridge or any time prior to theintroduction of the sample fluid, it can be appreciated that accordingto alternative embodiments, the substance may be introduced into thechamber together with or after introducing the sample fluid. In othercases, wherein the substance is composed of a combination ofconstituents or wherein the substance is the outcome of a chemicalreaction between more than one constituents, it is possible that atleast one constituent is pre-loaded while at least one other constituentis introduced with or after introduction of the sample fluid.

In case the chamber 303 is loaded with a substance, whether pre-loadedor loaded with/after introduction of the sample fluid, the procedureaffecting the sample fluid may be mixing of the sample fluid with thesubstance. Normally with reference to such a procedure, the sample fluidand the substance must be mixed thoroughly as lack of homogeneity willadversely affect subsequent analysis. According to certain embodimentsof the invention, in order to enable mixing, at least part (a portion)of the surface of the chamber, comprises a pressable portion made of anelastic polymer, for example polyurethane or silicone, or of a differentelastic material. Due to deformation (such as constriction) of thechamber, affected by pressing and/or releasing the pressable portion,fluid contained within the chamber will form a jet flow inside thechamber, a form of flow that enhances mixing. Hence, according toembodiments of the invention, it is possible to achieve mixing byalternatively pressing and releasing the pressable portion of thechamber. When the pressable portion is pressed the fluid flows away andwhen it is released it flows back, that is, the fluid flows back andforth.

In certain embodiments of the invention a pressable portion constitutesa part of a chamber's surface, for example, an upper surface of achamber or a certain percentage of its surface, while in differentembodiments of the invention the entire chamber is pressable. That is,100% of the chamber surface are also considered as a portion thereof.

Apart of or in addition to mixing, procedures affecting the sample fluidperformed in the chamber may be reactions that may occur between thesubstance and the sample fluid. The reaction may be a chemical reaction,for example oxidation/reduction, or a biochemical reaction such asbinding antibodies to ligands. The procedure may lead to changes inphysical and/or chemical states of the sample fluid or of cellscontained within the sample fluid. For example, it may affect changes inviscoelastic properties or in pH of the sample fluid; Concentration ofcells contained in a sample fluid may decrease due to dilution; Acellular membrane may become permeable enabling binding of coloringagents or antibodies contained within the substance to cellularcomponents, such as cytoplasmic granules; An oxidation or reduction ofdifferent cellular components may happen, such as oxidation ofhemoglobin contained in the red blood cells into methemoglobin; etc.

After the procedure has been completed, the resulting output fluid isreleased from the chamber. The releasing may be affected by positivepressure, “pushing” the fluid out of the chamber, for example if fluidis being pushed out of the chamber by pressing, or it may be affected bynegative pressure, for example if fluid is driven out of the chamber byphysical forces the “pull” it out, such as gravitational force or due toapplication of external forces such as vacuum. For example, in certainembodiments of the invention the flow of the output fluid from thechamber via the second opening into the analyzing compartment is drivenby a suction force generated by the vacuum pump 104 coupled to theanalyzing compartment, as will be described further with reference toFIG. 10.

It was mentioned before that the chamber is enclosed between two seals,wherein the preceding seal 306 prevents fluid from flowing out of thechamber via the first opening 301 while the succeeding seal 307 preventsfluid from flowing out of the chamber via the second opening. It shouldbe appreciated that prior to introduction of the sample fluid the twoseals 306 and 307 should prevent release of the substance from thechamber; then they should prevent release of the substance and/or thesample fluid during the procedure; and they should also preventunintentional release of the output fluid prior to the intentionalrelease thereof.

Due to simplicity considerations attention is drawn first to thesucceeding seal 307. It should be appreciated that breaking, orbreaching the succeeding seal allows output fluid flowing out of thechamber towards the second opening. According to certain embodiments,after breaching the seal it is not required anymore and therefore it maybe left open. Thus, the second seal 307 constitutes a “frangible seal”.It is possible to form the seal, e.g., of adhesive configured to be tobe broken by application of pressure exceeding a certain threshold.Applying excessive pressure on the pressable part of a chamber, whoseoutcome is super-threshold pressure at the position of the seal,breaches the succeeding seal. The output fluid is then released to flowthrough the second channel via the second opening into the analyzingcompartment. In other words, the output flow is conveyed to theanalyzing compartment via the second channel and via the second opening.

It is noted that mixing of the sample fluid with the substance byintermittently pressing the pressable portion of the chamber does notresult in super-threshold pressure at the position of the seal, whichtherefore remains intact. Alternatively or additionally, it is possibleto protect the seal from being affected be super-threshold pressure byapplying pressure on the channel between the chamber and the seal, henceobtaining a physical obstacle preventing pressure arising in the chamberto reach the seal. According to a different alternative it is possibleto apply the pressure further to the seal. According to this embodimentsupper-threshold pressure may reach the seal and breach it, however thephysical obstacle affected on the channel will prevent fluid fromflowing, until the obstacle is removed. It is further clarified, withreference to the two last alternative embodiments, that the pressureapplied on the channel in these cases is impermanent.

Further to understanding how breaching of the succeeding seal canhappen, attention is now drawn to the preceding seal 306. This seal hastwo different roles. The first role is preventing release of thesubstance from the chamber prior to the introduction of the samplefluid. However, when introducing the sample fluid, the preceding sealmust be broken, in order to allow such introduction. Yet, it waspreviously explained that in order to allow mixing using pressureaffected to the pressable portion of the chamber, the chamber must besealed from both sides. Therefore, the preceding seal has a second role:unlike the succeeding seal, the preceding seal must be re-sealed afterintroduction of the sample fluid, in order to allow mixing and in orderto prevent unintentional release of the output fluid from the chamber.

It was previously explained that the sample fluid may be introduced viathe first opening using a carrier. In light of the latter explanationrelating to re-sealing, it should be appreciated that in thoseembodiments wherein the carrier is left in the cartridge further to thesample fluid introduction, re-sealing should seal passage of fluid viathe gap existing between the carrier and the first channel's internalsurface.

FIGS. 4A and 4B depict a preceding seal 306, according to certainembodiments of the invention. The embodiments illustrated by the figuresare adapted for a carrier that remains inside the first channel furtherto the delivery, or introduction of the sample fluid.

In accordance with the illustrated embodiments, the depicted precedingseal 306 is comprised of two separate seals, namely, a first seal 401and a second seal 402. FIG. 4A depicts the preceding seal prior tointroduction of the sample fluid using a carrier 403, while FIG. 4Bdepicts the seal when the carrier is inserted, penetrating the precedingseal 306.

The first seal 401 is configured to prevent flow from the chamber viathe first opening prior to introduction of the sample fluid (the firstrole mentioned above). Hence, similar to the succeeding seal, the firstseal 401 may be a frangible seal, formed of adhesive or a plug. Uponinsertion of the carrier 403 into the chamber via the first opening, thecarrier 403 breaks seal 401, as illustrated in FIG. 4B.

The second seal 402 is in charge of re-sealing the chamber further tothe insertion of the carrier (the second role mentioned above). Thesecond seal is configured to prevent the leakage through the interfacebetween the carrier, more accurately, the outer surface of the carrier,and the inner surface of the channel. According to certain embodiments,the seal is comprised of a flexible ring mounted inside the channel (ano-ring). The inner diameter of the ring is smaller than the diameter ofthe carrier, hence, the ring allows the carrier to pass through, whileclosing tight around to prevent leakage. According to alternativeembodiments the first seal 401 and the second seal 402 may be swapped,that is, seal 402 may appear prior to the first seal 401.

Prior to continuing to additional and/or alternative embodiments, it isnoted that the carrier may be hollow inside. Hence, after the insertionthereof flow, or leakage out of the chamber may occur also through theinner space of the carrier. According to certain embodiments,illustrated and described, e.g., with reference to FIG. 14 below, thisleakage is prevented by a hydrophobic membrane located inside thecarrier.

FIGS. 5A and 5B depict a preceding seal, alternative to the sealillustrated in FIGS. 4A and 4B, according to certain embodiments of theinvention. Unlike the embodiment of FIGS. 4A and 4B wherein the seal 306is comprised of two seals (namely, these are the first 401 and second402 seals), the seal in the present figure is comprised of a singlemember whose functionality is similar to the functionality of seals 401and 402 when combined. In FIG. 5A, a stopper 501 with centeringshoulders is molded inside the first channel 302. Stopper 501 preventsflow from the chamber via the first opening 301, prior to theintroduction of the sample fluid. Upon insertion of a carrier 403, asillustrated by FIG. 5B, the center of the stopper 501 is breached, whilethe shoulders of the stopper block the interface between the outersurface of the carrier and the inner surface of the channel, preventingleakage further to the sample fluid introduction. According to certainembodiments stopper 501, that forms preceding seal 306, may be mad of asoft adhesive elastomer.

FIGS. 6A and 6B depict another alternative preceding seal, according tocertain embodiments of the invention. Similar to seal 501 of FIGS. 5Aand 5B, the present embodiments' seal 601 also describes a single sealcombining the functionality of the first and second seals (404 and 402)illustrated in FIGS. 4A and 4B. Unlike the stopper 501 (of FIG. 5) thatis configured for being breached by the carrier, according to thepresent embodiments the preceding seal is an enjected eyelet 601 with anintegrated plug 602, configured for being displaced, pushed by thecarrier. The eyelet 601 and the plug 602 may comprise different units ormay belong to the same unit, i.e., they me be coupled or not. Asillustrated in FIG. 6A, the plug is coupled to the eyelet and hence theyform the same unit. However, this is non-mandatory. The plug may becoupled, e.g., to the chamber or to the channel, or it may have nocoupling mechanism where applicable.

According to FIG. 6A, prior to the introduction of the sample fluid, theplug is closed, hence it prevents flow from the chamber via the firstopening. FIG. 6B illustrates introduction of sample fluid to the chamberwhile using a carrier such as a capillary. Upon insertion of thecarrier, the plug is pushed inwards, thus opening the channel, howeverthe eyelet 601 seals the interface between the outer surface of thecarrier and the inner surface of the channel, preventing leakagethereby.

Further to presenting several embodiments for applying a preceding seal,it should be appreciated that there may exist other embodiments, whereinfollowing the introduction, or delivery of a sample fluid into thechamber, the carrier is withdrawn from the first channel. An example forsuch a carrier is a needle attached to a syringe usable to deliver thesample fluid into the first chamber. In such cases the preceding sealhas to re-seal the opening left after the carried is withdrawn. Anexample for such a seal is a known per se septum.

Before describing the invention further, a summary of a process ofpreparation of a sample fluid for analysis, according to certainembodiments of the invention is presented. A carrier 403 of a samplefluid is inserted via the first opening 301 into the first channel 302.The carrier breaches the preceding seal 306 coupled to the first channeland delivers the sample fluid into the chamber 303. Inside the chamber aprocedure is affected to the sample fluid, such as mixing the deliveredsample fluid with a substance pre-loaded to the chamber, thus obtainingan output fluid. Mixing is enabled by applying an intermittent pressureon a pressable portion of the chamber. Upon completion of the procedure,the succeeding seal 307 becomes broken by pressing the chamber in a waygenerating a super-threshold pressure at the position of the succeedingseal, resulting in a release of the obtained output fluid from thechamber. The released output fluid then flows via the second channel 304and the second opening 305 into the analyzing compartment 203, whereinit is subjected to analysis.

FIG. 7 presents a cartridge comprising a chamber (e.g., a mixingchamber) containing two compartments, according to certain embodimentsof the invention. The two compartments 701, which may be pre-loaded witha substance, are interconnected by a narrowing 702. The firstcompartment is coupled to the first opening 301 via a first channel 302,while the second compartment is coupled to the second opening 305 via asecond channel 304. At least one, and possibly two of the compartmentsincludes a pressable portion.

In case that both compartments have pressable portions, it is possibleto achieve mixing by alternating pressure applied to the two pressableportions (each compartment in a time). The narrowing 702 between thecompartments 701 causes jet flow, hence enhancing mixing. Breaking thesucceeding seal 308 may be caused, e.g., by simultaneously pressing bothcompartments and/or by applying stronger pressure than the pressureapplied for mixing.

In case that there is only one pressable portion, on one of thecompartments, it is possible to achieve mixing by intermittentlypressing this portion. Breaking the succeeding seal 308 may be caused byapplying excessive pressure on the pressable portion.

In the description of the latter embodiment it was written that thesubstance is pre-loaded into the two compartments. Prior to continuingto describe additional and alternative embodiments, it should be notedthat the substance may be loaded into only one compartment, ifapplicable.

In addition, instead of having two compartments having the formillustrated in FIG. 7, it is possible to have alternative embodiments,having other forms. For example, the chamber may appear from the outsideas the chamber illustrated in FIG. 3, having a partitioning memberinside. An opening or even a valve in the partitioning member mayfunction as the narrowing in FIG. 7.

In the embodiments described so far the cartridge comprised one chamber.However, the invention is not limited thereby, and, in differentembodiments the cartridge may contain more than one chamber, wherein thechambers are connected in series. Hereinafter, one or more chambersenclosed between frangible seals and connected in series constitute a“preparation unit”. Therefore the cartridge described, e.g., withrespect of FIG. 3 can be defined as a cartridge comprising onepreparation unit containing a single chamber. Similarly, the cartridgeof FIG. 7 is also a cartridge comprising one preparation unit containinga single chamber.

FIG. 8 presents a cartridge comprising a preparation unit composed oftwo chambers, according to certain embodiments of the invention. A firstchamber 801 (e.g., a first reagent chamber), coupled to a first opening301, is a pressable chamber, while a last chamber 802 (e.g., a secondreagent chamber), coupled to a second opening 305, might be eitherpressable or not pressable. The two chambers, are connected by aconnecting channel 803 sealed by a seal 804. The two chambers areenclosed, each, between seals, whereas the first chamber 801, ispreceded by a preceding seal 306 and the last chamber 802, is succeededby a succeeding seal 307. Seal 804 is a succeeding seal with respect tochamber 801, while with respect to chamber 802 it is a preceding seal.

In the illustrated case, the first and the second openings of apreparation unit constitute respectively the first and the secondopenings of a cartridge. This however is non-limiting, and in differentembodiments, for example in the embodiments described by FIG. 9 below,the first and the second openings of a preparation unit may be distinctfrom the first and the second openings of a cartridge.

While each chamber has a respective input fluid and a respective outputfluid, the input fluid of the first chamber, introduced thereto via thefirst opening, is a sample fluid. Inside the first chamber a procedureaffecting the fluid is performed. This procedure is referred to as a“first procedure”. In case the procedure includes mixing, it isperformed as described with reference to FIG. 3. By affectingappropriate pressure on seal 804 it may be breached (see FIG. 3 and thedescription relating thereto), resulting in release of the output fluidfrom the first chamber, conveying it to the last chamber. The outputfluid of the first chamber becomes, therefore, an input fluid of a lastchamber.

At this stage it should be considered that if seal 804 is a frangibleseal, further to breaching it the path between the two chambers 801 and802 is left open and flow is possible at both directions, that is from801 to 802 and from 802 to 801. Hence, the two chambers form, in effect,two compartments of a single chamber. Therefore, in embodiments having afrangible seal in the connecting channel 803, further to breaching thisseal the output fluid of the first chamber 801 can flow back and forthbetween the two former chambers, while being affected by the procedureof the last chamber. That is, the two chambers form two compartments ofthe last chamber. The description provided with reference to FIG. 7 maybe applicable to this last chamber comprised of two compartments (801,802).

Understanding this, it is noted that because 802 and 801 now effectivelyform a single chamber, the channel connecting this single chamber to thesecond opening, that is the second channel, may couple “compartment” 801with the opening instead of “compartment” 802.

However, if seal 804 is re-sealable, further to conveying chamber 801'soutput fluid to chamber 802, the latter chamber can be re-sealed,therefore the description provided with reference to chamber 303 in FIG.3 may be applicable thereto. An example of a re-sealable seal is avalve. Moreover, instead of using a sealable seal, certain embodimentsmay have a re-sealable connecting channel 803, while re-sealing may beperformed, for example, by applying pressure to the connecting channel803, hence obtaining a physical obstacle preventing fluid from flowingat this route.

Inside the last chamber a procedure, referred to as a “secondprocedure”, is performed. By affecting appropriate pressure on thesucceeding seal 307, it may be breached, thus resulting in release ofthe output fluid from the last chamber towards the second opening 305.The output fluid of the last chamber constitutes, therefore, an outputfluid of a preparation unit. The output fluid of the preparation unitflows via the second opening 305 into the analyzing compartment 203,wherein it is subjected to analysis.

After describing the embodiments of FIG. 8 with reference to FIGS. 3 and7, it should be appreciated that these embodiments are non-limiting as apreparation unit may be comprised of one, two, or more than twochambers. In general, a preparation unit may be comprised of one or morechambers connected in series, each chamber is enclosed between frangibleseals. Each chamber is configured for receiving an input fluid,performing a procedure affecting the fluid thereby generating an outputfluid, and releasing the output fluid. A first chamber of the one ormore chambers is coupled to a first opening, while a last chamber iscoupled to a second opening. A first chamber is a pressable chamber,whereas the preparation unit may include additional pressable chambers.The input fluid of the first chamber is a sample fluid while the inputfluid of each one of the other chambers is the output fluid of thechamber preceding thereto. The output fluid of the last chambercomprises the output fluid of the preparation unit to be subjected toanalysis.

It is noted that according to certain embodiments in a preparation unitincluding, e.g., two chambers, it is possible to apply pressure on thefirst chamber in order to breach the seal inbetween. Alternatively, theseal may be breached by applying pressure on the second chamber.

Further to understanding how the preparation of FIG. 8 operates, itshould be appreciated that other embodiments may have preparation unitswith more than two chambers, wherein each chamber is enclosed betweenseals, namely a preceding seal and a succeeding seal. The precedingand/or the succeeding seals may be frangible or re-sealable. That is,according to certain embodiments of the invention, there may existseries of two or more chambers. Amongst the serialized chambers thereare, at least, a first chamber and a last chamber, while the firstchamber is coupled to the first opening (a sample fluid is introducedtherethrough into the first chamber) via a first channel and the lastchamber is coupled to the second opening via the second channel.

Each chamber in the series is configured to perform a procedure. Hence,if the first chamber obtains the sample fluid, it may be appreciatedthat the procedure respective of the first chamber affects this samplefluid, yielding a derivative of the sample fluid. The derivativemanifests a change having occurred in a sample fluid or in cellscontained within the sample fluid. The occurred change may be a chemicalchange, a biochemical change or a physical change. Examples of achemical change are change in pH, oxidation/reduction of cellularcomponents or hinging of chemical agents, such as dyes thereto; examplesof a biochemical change is binding of antibodies to ligands; whereasexamples of physical change are changes in viscoelastic properties, intemperature or in concentration of diluents. According to an alternativeview, the sample fluid may be considered as a derivative of itself,i.e., a derivative of the sample fluid. Hence, the procedure obtains asinput a derivative of the sample fluid, and yields an output which is aderivative of the derivative. In order to clarify this, the derivativesare given “names”: the chamber obtains a first derivative of the samplefluid, while the procedure's output, and hence also the chamber'soutput, is a second derivative of the sample fluid.

Understanding how the first chamber operates, it should be appreciatedthat this is correct to all the other chambers in the series: eachchamber obtains an input fluid which is a first derivative of the samplefluid, the first derivative is the input of the procedure operative withrespect of the chamber. Then, the output of the procedure is a secondderivative of the sample fluid, while this second derivative is also theoutput of the chamber.

Because the chambers are consecutive, so are the procedures: theprocedure of a certain cell in the series yields a second derivative ofthe sample fluid, which is the output of the chamber. The consecutivechamber obtains the second derivative being output of the precedingchamber, while here (in the consecutive chamber) it is considered as afirst derivative, being input to the consecutive chamber's procedure.The output of the consecutive chamber's procedure is a respective secondderivative of the sample fluid. This second derivative is furtherconveyed to the coming chamber, a chain that lasts until the lastchamber conveys its respective second derivative of the sample fluidtowards the second opening.

An example for consecutive procedures is an immune-labeling of cells:labeling with a primary antibody is performed in a first chamberfollowed by a consecutive labeling with a secondary antibody, performedin a last chamber. Another example is differential staining of whiteblood cells of a blood sample, with two staining reagents, that must beseparated during storage. Procedure of staining with a first reagent,performed in a first chamber, is followed by staining with a secondreagent, performed in a consecutive, possibly last chamber.

It should be appreciated that in accordance with embodiments of thepresent invention the procedure is performed inside the chambers,wherein each chamber adds a stage in the preparation of the outputfluid, all together resulting in a cumulative continuous process. Thisis unlike having the procedures performed in a dedicated chamber. Hence,efficient and complete mixing of the fluid and the reagents is affected.

FIGS. 9A and 9B presents two configurations of a cartridge comprisingtwo preparation units, according to certain embodiments of theinvention. One of the preparation units comprises a single chambercontaining two interconnected compartments 701. This preparation unithas been described above, with reference to FIG. 7. The otherpreparation unit comprises two chambers 801 and 802, connected by achannel 803 and sealed by a seal 804. This preparation unit has beendescribed above, with reference to FIG. 8. Each preparation unit has arespective first opening 301 and a respective second opening 305. Thefirst openings of both preparation units constitute the first openingsof the cartridge.

The two configurations of the cartridge, depicted by FIGS. 9A and 9B,differ respective to number of the cartridge second openings presenttherein. The cartridge depicted at FIG. 9A has a single cartridge secondopening 901 which is distinct from the second openings 305 of thepreparation units. The cartridge depicted at FIG. 9B has two preparationunit's second openings 305, which, in this case, constitute also secondopenings of the cartridge.

In the described embodiments each preparation unit of a cartridge isconfigured for introduction of a sample fluid by a respective carrier.This however is not limiting, and in certain embodiments preparationunits of a cartridge may be configured for introduction of sample fluidfrom a single carrier. The sample fluid may be introduced into thepreparation units simultaneously or consequently, as will be explainedfurther below with reference to FIG. 14.

The output fluid of each preparation unit may flow into the analyzingcompartment at different timings and may be subjected to separateanalysis, as will be explained further below with reference to FIGS. 13Band 13B.

Existence of two parallel preparation units enables performing twoseparate independent procedures affecting the sample fluid. For example,in certain embodiments of the invention, the cartridge is configured forperforming a complete blood count. The cartridge comprises two parallelpreparation units, whereas one preparation unit is configured forpreparation of red blood cells for analysis, while the other preparationunit is configured for preparation of white blood cells for analysis(the abovementioned procedures will be explained in details below, withreference to FIGS. 13A and 13B).

Although the cartridges depicted by FIGS. 9A and 9B comprise twopreparation units, this is not limiting. The number of preparation unitsconstituting a cartridge, as well as the number of chambers constitutingeach preparation unit and the number of chambers containing more thanone compartment may differ, as configuration of a cartridge is tailoredfor performance of desired procedures and/or for purpose of preparingthe sample fluid for certain analysis procedures.

FIG. 10 schematically illustrates an analyzing compartment, according tocertain embodiments of the invention. The analyzing compartmentcomprises an analysis vessel 1002, configured for receiving the outputfluid conveyed by preparation unit or units and for presenting it in away allowing analysis, and from a third channel 1004, coupled to theanalysis vessel and configured for emptying disposable output fluidtherefrom. The analysis vessel and the third channel together comprisean analyzing unit. A waste container 1005, configured for storingdisposed output fluid is coupled to the analysis unit via the thirdchannel 1004. The waste container 1005 is also coupled to a vacuum pump104 via a fourth channel 1006.

An output fluid flows from a preparation unit into the analyzing unitvia a third opening 1001. Inside the analysis vessel 1002 the outputfluid is presented to an analyzing system 101. After being subjected toanalysis the output fluid is disposed via the third channel 1004 intothe waste container 1005 and stored therein.

The flow of the output fluid inside the analyzing unit is driven by asuction force generated by the vacuum pump 104, which may be a part ofthe analyzing system 101. The vacuum pump is coupleable to the analyzingunit through a fourth opening in the analyzing compartment, connectedvia the fourth channel 1006 to the fifth opening 1008 in the wastecontainer 1005. Although the suction force is applied to the wastecontainer, the stored output fluid does not flow out therefrom, becausethe waste container should be designed as a liquids trap in any wayknown per se. In the figure the fifth opening 1008 is located above thelevel of the stored output fluid, hence schematically representing sucha liquid trap.

In certain embodiments of the invention, illustrated by FIG. 10, ananalysis vessel of the compartment is a microchannel 1003 configured toalign cells contained in the output fluid into a single plane, allowingtaking images of the flowing cells by a camera 107, or probing by afocused light beam/laser beam as done in a cytometer. The aligning ofthe cells may be performed by a method known as viscoelastic focusing.Viscoelastic focusing is described in PCT Publication No. WO2008/149365entitled “Systems and Methods for Focusing Particles”, while amicrochannel configured for viscoelastic focusing is further describedin PCT Publication WO2010/013238, entitled “Microfluidic System andMethod for Manufacturing the Same”. The aligned cells may then beoptically analyzed, through a transparent or translucent surface of themicrochannel.

FIG. 11 schematically illustrates an alternative analyzing compartment,configured for determination of blood hemoglobin level, according tocertain embodiments of the invention. This compartment too includes ananalyzing unit comprises an analysis vessel 1002 which consists of ananalyzing chamber 1101 coupled to a long small cross sectioned thirdchannel 1103.

The analyzing chamber 1101 may contain a powdered oxidizing agent and/ora lysing agent. The agent may be Sodium Dodecyl Sulfate (SDS), TritonXor another oxidizing/lysing agent suitable for the case. When thechamber is filled with the output fluid, which in this case is aderivative of a blood sample, the oxidizing agent becomes dissolved. Thedissolved oxidizing agent lyses the red blood cells of the derivative ofthe blood sample leading to release of hemoglobin. The releasedhemoglobin is then oxidated by the oxidizing agent to methemoglobin(which is a form of hemoglobin which cannot release bound oxygen).Concentration of methemoglobin is then determined using a spectrometer,by measuring an absorption of one or more wavelengths. That is, theanalyzing module 105 of system 101 (see FIG. 1) should comprise aspectrometer, in this case.

According to certain embodiments, powdered agent may freely resideinside chamber 1101. Alternatively, it may coat the inner surface of thechamber 1101. For the purpose of enlarging the contact area between theagent and the derivative of the blood sample, according to certainembodiments the inner surface of the chamber may contain projectionssuch as pillars, coated with the agent. Alternatively, for the samepurpose, a powdered oxidizing agent may be attached to a carrier, suchas sponge, filling the chamber.

Having understood this, it should be noted that a chamber containingcoated projections is not limited to powdered oxidizing and/or lysingagents and/or to blood samples. A chamber having projections coated witha powdered agent, or even with other forms of agents such as gel, may beused in other occasions and situations wherein enlarging the contactarea inside an analysis compartment's analysis vessel may be beneficial.

Those versed in the art would appreciate that the processes of celllysis, hemoglobin oxidation and measuring an absorption require acertain minimal time interval each, Therefore, the derivative of theblood sample must be retained inside the analyzing chamber. According toembodiments of the invention, it is possible to achieve retention byapplying high resistance to the flow, hence slowing it down. One way forapplying such high resistance is by means of a long third channel 1003having a small cross section coupled to the analyzing chamber 1101. Whenthe channel is empty, no resistance to flow is present, therefore thederivative of the blood sample flows freely into the analysis vessel andthe analyzing chamber via the third opening. However, filling the thirdchannel with a derivative of the blood sample causes the resistance toraise, leading to a nearly complete cessation of flow.

FIG. 12 schematically illustrates an analyzing compartment, comprisingtwo analyzing units, according to certain embodiments of the invention.One of the analytical units comprises a microchannel 1003, such as theanalyzing unit depicted in FIG. 10 and described with reference thereto.The other analyzing unit comprises an analyzing chamber 1101, such asthe analyzing unit depicted in FIG. 11 and described with referencethereto. According to such embodiments the two analyzing units may becoupled on one side to a third opening 1001 for purpose of obtaining theoutput fluid. On the other side they may be coupled to the wastecontainer 1005, wherein disposable fluid may be disposed. That is, thetwo analyzing units are coupled in parallel.

It is noted that such parallel coupled analyzing units within ananalyzing compartment enable to perform in parallel two separate typesof analysis of the output fluid. For example, using the analyticalcompartment depicted by FIG. 12 cell counting and measuring ofhemoglobin level of a derivative of a blood sample may be performed. Itis worth noting, that the two types of analysis are performed usingdifferent analyzing modules 105 in system 101 (see FIG. 1), e.g., acamera and a spectrometer.

FIGS. 13A and 13B schematically illustrate a cartridge comprising apreparation compartment and an analyzing compartment, according tocertain embodiments of the invention.

In has been stated above, with reference to FIG. 2 that in someembodiments the analyzing compartment 203 coupled to the cartridge 201,is not a part thereof, while in other embodiments the analyzingcompartment 203 may be considered as part of the cartridge. According tosuch embodiments 201 relates to a “preparation compartment” while acartridge 204 may be composed of a preparation compartment 201 and ananalyzing compartment 203. To this end, and due to simplicityconsideration, FIGS. 13A and 13B are presented and described wherein acartridge 204 has a preparation compartment 201 and an analysiscompartment 203. However, this is non-limiting and the descriptionapplies also to the other convention, wherein 201 is the cartridge.

Two different embodiments of cartridge 204 are depicted by FIGS. 13A and13B. First, the description of features common to both configurations isbrought, and then, dissimilarities therebetween shat be discussed.

The preparation compartment 201 of the cartridge 204 has been describedin detail above, with reference to FIGS. 9A and 9B. In the examplepresented in FIGS. 13A and 13B, the preparation compartment comprisestwo preparation units, the first unit and the second unit. The firstpreparation unit, comprising a single chamber (e.g., a mixing chamber)containing two interconnected compartments 701, has been described indetail above, with reference to FIG. 7. The second preparation unit,comprising two chambers 801 (e.g., a first reagent chamber) and 802(e.g., a second reagent chamber), has been described in detail above,with reference to FIG. 8.

The analyzing compartment 203 of a cartridge 204 has been described indetail above, with reference to FIG. 12. The analyzing compartmentcontains two analyzing units. One of the analyzing units, comprising amicrochannel 1003, is configured to align cells contained in the outputfluid into a single plane allowing taking images of the flowing cellsusing a camera, or probed by a focused light beam/laser beam as done ina cytometer. This analysing unit has been described in detail above,with reference to FIG. 10. The other analyzing unit, comprising ananalyzing chamber 1101 coupled to a long small cross-sectioned thirdchannel 1004, is configured for determination of hemoglobin level, e.g.,using a spectrometer. This analysing unit has been described in detailabove, with reference to FIG. 11.

To allow flow of the output fluid prepared for analysis from thepreparation compartment to the analysis compartment the two compartmentsare interconnected by means of the second opening of the preparationcompartment coupled to the third opening of the analyzing compartment.The two configurations of the cartridge, depicted by FIGS. 9A and 9B,differ respective to a number and position of the second openings of thepreparation compartment and correspondingly the coupled thereto thirdopenings of the analyzing compartment. Thus, in the cartridge depictedby FIG. 13A there is a single second opening 901 of preparationcompartment 201, connected to the second openings 305 of bothpreparation units. The single second opening 901 of the preparation unitis coupled to a single third opening 1001 of the analyzing compartment.In contrast, in the cartridge depicted at FIG. 13B the second openings305 of both preparation units form also second openings of thepreparation unit, and they are directly coupled to the respective twothird openings 1001 of the analyzing compartment.

According to certain embodiments of the invention the cartridge 204 isconfigured to allow a performance of blood count, while a sample fluidintroduced into the cartridge is a blood sample. A blood count performedby the cartridge may include determination of number of red blood cells,white blood cells (total count) and platelets present in the sample, aswell as determination of number of each one the of white blood celltypes (differential count). The white blood cell types may beneutrophils, lymphocytes, monocytes, eosinophils and monocytes or partthereof. Those versed in the art would appreciate that there areadditional types and sub-types of white blood cells, and therefore theinvention is not limited to the types mentioned. Furthermore, theinvention is not limited to the blood cells mentioned and it may beapplicable to any type of cells circulating in the blood, including,e.g., circulating tumor cells, platelets aggregates and others.

Attention is drawn now to describing in detail the processes ofpreparation of a blood sample for analysis, whereas the analysis is ablood count.

In the described embodiments of the invention, cells counting may beperformed by means of acquiring images of flowing cells by a camera orby probing by a focused light beam/laser beam as done in a cytometer. Inorder to allow reliable counting, the cells mast be brought into thefocal place of the optics. Hence, the cells should be aligned in asingle plane, e.g., by viscoelastic focusing. The method is based onsuspending cells in a focusing medium of certain viscoelastic propertiescausing the cells suspended therein to align into a single plane ifbeing flowed in a microchannel of a certain geometry. Thereforepreparing a sample fluid for counting, performed in preparationcompartment 201 of a cartridge 204, includes adding focusing media tothe sample fluid, thus yielding a derivative of the sample fluid. Thishowever is not-limiting and in different embodiments cells may bealigned using other methods, whereas preparing a sample fluid forcounting, performed in a preparation unit of a cartridge, may includedifferent procedures.

The first preparation unit is configured for preparing a blood samplefor determination of number of red blood cells, white blood cells (totalcount) and platelets present therewithin. A substance contained inchamber 701 comprises focusing medium with addition of surfactants. Thefocusing medium comprises a buffer containing soluble high molecularweight polymers. The buffer may be any isotonic buffer suitable formanaging living cells, for example it may be Phosphate Buffered Saline(PBS). Examples of soluble polymers suitable for providing the bloodsample with viscoelastic properties are polyacrylamide (PAA),polyethyleneglycol (PEG), Propylene Glycol, etc. The surfactants addedto a focusing media act as sphering agents, i.e., they cause the shapeof red blood cells to change from biconcaive discs into spheres,allowing to obtain better images of cells. Examples of surfactants areSDS (Sodium Dodecyl Sylphate) and DDAPS(dodecyldimethylammoniopropanesulfonate). The composition of thefocusing medium is disclosed, e.g., in PCT Publication No. WO2008/149365entitled “Systems and Methods for Focusing Particles”.

The procedure performed by chamber 701 is mixing of the delivered bloodsample with a focusing medium. After mixing has been completed thesucceeding seal 305 becomes breached by pressure, allowing the generatedoutput fluid to flow into the analytical compartment.

The second preparation unit is configured for preparing a blood samplefor differential count of white blood cell types. In certain embodimentsof the invention the preparation includes chemical staining of cells,whereas two consecutive staining procedures are performed in chambers801 and 802 of the preparation unit.

The substance contained in chamber 801 may comprise cell stainingreagents dissolved in the focusing medium. Examples for cell stainingreagents are Phloxine B, Biebrich Scarlet and Basic Orange 21. As afixation of cells may be needed in some cases, fixating reagents, forexample formaldehyde or formalin, may also be contained. Followingmixing of the blood sample with the substance, an incubation may beperformed, allowing staining. Upon expiration of a predeterminedincubation time a seal 804 separating the chamber 801 from the chamber802 becomes breached by pressure, resulting in release of the generatedoutput fluid towards the chamber 802.

The substance contained in chamber 802 may comprise other cell stainingreagents dissolved in the focusing medium. Examples for cell stainingreagents are Methyl Green, Methylene Blue and Borrel's Blue. Followingmixing of an input fluid (which constitutes the output fluid of chamber801) with a substance, a second incubation may be performed, allowingthe second staining process to occur. Upon expiration of a secondpredetermined incubation time the succeeding seal 307 of the secondpreparation unit becomes breached by pressure allowing generated outputfluid to flow into the analytical compartment.

In other embodiments of the invention preparing cells for analysisincludes immuno-based staining of the cells. In these embodiments one orboth chambers of the preparation unit contain reagents suitable forimmune-staining, whereas the reagents and the focusing medium may becontained within a single chamber or in different chambers. Examples ofreagents suitable for immune-staining are antibody-coated micro beads ofdifferent colors, such as CD14/CD15 and a combination of stains.

Attention is turned now to a detailed description of the process ofpresentation for analysis of the output fluid of the preparation units,whereas the analysis is blood count.

The output fluids flowing out of the second openings 305 of bothpreparation units are conveyed to a single channel that is coupled tothe analysis vessels of both analyzing units. Analysis of the outputfluids is performed sequentially. The sequentially analysis is enabledby temporal separation of the two output fluids, a separation controlledin the preparation compartment. As described above the preparationprocess performed by a first preparation unit includes mixing in asingle chamber without incubation, while the preparation processperformed by a second preparation unit includes, in addition to mixingin two different chambers, two staining procedures that might requireincubation time. Hence, the output fluid of the first preparation unitis ready to flow into the analyzing compartment before the output fluidof the second preparation unit is ready.

Upon flowing into the analyzing compartment 203 the output fluid of thefirst preparation unit is divided between the two illustrated analyzingunits. Part of the fluid enters the microchannel 1003, wherein the cellswithin the output fluid become aligned into a single plane in a wayexplained with reference to FIG. 10. The aligned cells may then beoptically analyzed, through a transparent or translucent surface of themicrochannel. The output fluid then flows into waste container 1003,wherein it is stored.

The other part of the output fluid enters the analyzing chamber 1101,wherein the cells within the output fluid become lyzed and theirhemoglobin content quantified in a way described in reference to FIG.11.

The flow of the output fluid of the first preparation unit into theanalyzing compartment must be aborted prior to breaching the succeedingseal 307 of the second preparation unit, to prevent mixing of the outputfluids that will hamper analysis. This is enabled due the second channel304 of the first preparation unit being re-sealable. The re-sealing ofthe channel may be performed, for example, by pressure applied to thesucceeding seal or to another area of the second channel.

The long and small cross-sectioned third channel 1103 coupled to chamber1101, enhances resistance to flow at the chamber. Hence, upon breachingsucceeding seal 307 of the second preparation unit, the substantiallyall the output fluid flows into the analyzing compartment and conveyedto the microchannel 1003 Instead of being split between the two analysisunits. Inside the microchannel 1003 the cells within the output fluid ofthe second preparation unit become aligned into a single plane henceallowing optical analyzed. The output fluid then flows into the wastecontainer, wherein it is stored.

It is further noted that in FIGS. 10-13B, the waste container 1005appears to be in the analyzing compartment 203. Yet, this isnon-mandatory. It was previously explained, with reference to FIG. 2,that the analyzing compartment 203 may be positioned in a gap, or awindow, inside the cartridge. In such embodiments the cartridge maycomprise the waste container, wherein in such embodiments an opening inthe third channel 1004 would be used for interfacing the third channelin the analyzing module with the waste container in the cartridge orwith a channel leading thereto.

Having understood the structure and functionality of a cartridge, inaccordance with certain embodiments of the invention, a sampler is nowdescribed. FIGS. 14A, 14B and 14C, schematically depict samplers,according to certain embodiments of the invention. A sampler 1400 isconfigured to sample fluid and to introduce it into the cartridge 204.The sampler depicted by 14A comprises a carrier 1401, attached to ahandle 1402. In the described non-limiting embodiment the carrier is acapillary. Inside the capillary a hydrophobic membrane 1404 is affixedin a pre-determined distance from the capillary outlet. The capillary1401 might be any type of capillary with a hydrophobic membrane affixedinside and suitable for the case, for example capillaries manufacturedby DRUMMOND Aqua-Cap™ Microdispenser.

It is noted though that the hydrophobic membrane 1404 does not limit theinvention and other mechanisms are allowed for ensuring that the amountof sample fluid in the capillary is precise. For example: it is possibleto use a shorter capillary whose volume is the precise required volume.Alternatively, another constriction element can be used instead of themembrane, such as an orifice.

Fluid sampling is performed by immersing the outlet of the capillary1401 in the fluid. A person versed in the art would appreciate that thefluid is driven into the capillary by capillary force. The hydrophobicmembrane 1404 affixed inside the capillary 1401 does not interfere withthe process, as it allows the air displaced by a driven inside fluid toflow out. The fluid fills the capillary until reaching the hydrophobicmembrane. It should be appreciated that due to the hydrophobic nature ofmembrane 1404, the fluid does not come into contact therewith. Thereforthere is no sample fluid absorbance in the membrane, or in other words,no loss of fluid volume occurs in favor of the membrane. Hence the finalvolume of a sampled fluid is determined by a distance of the hydrophobicmembrane 1404 from the capillary outlet and by the capillary's diameter.

Once the fluid has been sampled it is delivered, introduced, into thecartridge 204 by inserting the capillary 1401 through the first opening301 thereof. At this stage only a limited leakage of a sample fluid fromthe capillary into a chamber 303 occurs, as the fluid is being heldinside by capillary forces. Next, a plunger 1405 is used to push thesampled blood out of the capillary into the chamber 303. The plunger1405, depicted in FIG. 14B comprises a plunging member 1406 attached toa holding member 1407. The plunging member 1406 is configured to beinserted into the capillary 1401 through a capillary inlet 1403 locatedin the handle 1402. The plunger pushes the hydrophobic membrane 1404until it reaches the capillary outlet, optionally resulting in thedelivery of the entire sample fluid into the chamber 303. It should beconsidered though that if the plunging member 1406 is not long enoughfor reaching the capillary outlet, a certain dose of fluid will remainin the capillary. Hence the volume of the sample fluid delivered intothe chamber is determined by a length of a plunging member 1406 of aplunger 1405. The capillary's diameter is known in advance, and so isthe length of the capillary as well as the length of the plunger. Hence,the volume of the fluid transferable by the sampler can bepredetermined.

It must be appreciated that the mechanisms of sampling and plungingdescribed above enable delivery into the chamber of a fixed volume ofsample fluid. The ability to deliver a fixed volume of a fluid is veryimportant, as any deviation in the delivered volume may affect thereliability of the sequential analysis. Moreover there is no need toflush the blood out of the carrier (in this case the capillary) as isthe case in other applications because the hydrophobic membranethoroughly dispenses all the blood into the first chamber.

With reference to certain embodiments, the plunger 1405 is a part ofanalyzing system 101, whereas the plunger is inserted into the cartridge204 upon placement thereof inside the cartridge holding unit 103 of ananalyzing system 101. However, in different embodiments the plunger mayconstitute a separate device, whereas the insertion of a plunger intothe cartridge may be performed prior to placement thereof into thecartridge holding unit 103.

In different embodiments, illustrated by FIG. 14C, the sampler iscomprised of two carriers 1401, wherein sampling of the fluid by thecarriers is performed sequentially. However, this is not limiting andembodiments in which sampling of the fluid by the carriers is performedsimultaneously may exist.

The sampler comprising two carriers may be used, such as the oneillustrated in FIG. 14C, for sampling and delivery of blood into acartridge configured to allow performance of blood count, such as thecartridge described above with reference to FIG. 13. Sometimes, the twocarriers of the sampler may comprise anticoagulant-coated capillarieswith a hydrophobic membrane. An anticoagulant, coating the capillaries,serves to prevent clotting of sampled blood. An example of ananticoagulant is EDTA (Ethylenediaminetetraacetic acid).

It must be appreciated that a fluid volume sampled by each carrier 1401of the sampler 1400 and delivered into the cartridge 204 may be as smallas 20 μl and possibly even less. Therefore, performance of a blood countusing the sampler 1400, the cartridge 204 and the analyzing system 101requires obtaining of as little as a single drop of blood from theindividual. Such a small volume of blood may be obtained by pricking thefingertip or forearm in a way performed for example by home bloodglucose monitoring devices, thus sparing driving blood from a vein,which is less convenient for patients, especially children.

The invention claimed is:
 1. A disposable cartridge, comprising: aninlet configured to receive a sample fluid that includes blood; a firstreagent chamber wherein at least a portion of a surface of the firstreagent chamber comprises a pressable portion; a first reagentpre-loaded and sealed within the first reagent chamber; a second reagentchamber wherein at least a portion of a surface of the second reagentchamber comprises a pressable portion; a second reagent pre-loaded andsealed within the second reagent chamber; a flow channel interconnectingthe first reagent chamber and the second reagent chamber, the flowchannel, the first reagent chamber, and the second reagent chamberdefining a flow path; wherein a first frangible seal prevents flowbetween the first reagent chamber and the second reagent chamber; thefirst frangible seal configured to open in response to pressure appliedby contents of one of the reagent chambers against the first frangibleseal; wherein the cartridge is configured so that when the firstfrangible seal opens, the first reagent contacts the second reagent; anda mixing chamber, wherein a fluid substance is pre-loaded and sealedwithin the mixing chamber.
 2. The disposable cartridge of claim 1,wherein the cartridge further comprises another frangible sealassociated with the mixing chamber, the another frangible sealconfigured to open in response to pressure above a predeterminedthreshold exerted on the mixing chamber.
 3. The disposable cartridge ofclaim 1, further comprising: the second reagent chamber being coupleableto an analyzing unit, which performs analysis of an output fluid; and asecond frangible seal preventing flow between the second reagent chamberand the analyzing unit, wherein the first and second frangible seals areconfigured to open at different times.
 4. The disposable cartridge ofclaim 1, wherein at least one of the first and second reagent chambersis coupleable to an analyzing unit, which performs analysis of an outputfluid.
 5. The disposable cartridge of claim 1, wherein a secondfrangible seal in the flow path is configured to prevent the first orsecond reagent from flowing out of the second reagent chamber when thesecond frangible seal is intact.
 6. The disposable cartridge of claim 1,wherein at least one of the first reagent and the second reagentincludes at least one high molecular weight polymer in fluid form. 7.The disposable cartridge of claim 1, wherein the first reagent chamberincludes an inlet seal configured to prevent the first reagent fromflowing via the inlet when the inlet seal is intact.
 8. The disposablecartridge of claim 7, wherein the inlet seal comprises a re-sealableseal.
 9. The disposable cartridge of claim 1, wherein at least one ofthe first and second reagent chambers is configured to convey an outputfluid to an analyzing unit via an opening.
 10. The disposable cartridgeof claim 9, wherein a second frangible seal in the flow path isconfigured to prevent the first or the second reagent from flowing viathe opening when the second frangible seal is intact.
 11. The disposablecartridge of claim 1, further comprising: at least one of the firstreagent chamber and the mixing chamber being provided with an entrysealing member, wherein the entry sealing member prevents at least oneof the first reagent and fluid substance from flowing out of the atleast one of the first reagent chamber and the mixing chamber via theinlet when the entry sealing member is intact, the entry sealing memberconfigured to be punctured by a capillary tube, and wherein the entrysealing member is configured to seal around a circumference of thecapillary tube that punctures the entry sealing member and is insertedinto the at least one of the first reagent chamber and the mixingchamber.
 12. The disposable cartridge of claim 11, wherein the entrysealing member comprises: an inlet seal and a capillary seal, whereinthe inlet seal prevents the at least one of the first reagent and fluidsubstance from flowing out of the at least one of the first reagentchamber and the mixing chamber via the inlet when the inlet seal isintact, the inlet seal configured to be punctured by the capillary tube,and wherein the capillary seal is configured to seal around thecircumference of the capillary tube inserted through the capillary sealand into the at least one of the first reagent chamber and the mixingchamber.
 13. The disposable cartridge of claim 11, wherein the mixingchamber comprises at least a first compartment and a second compartment,wherein a narrow passage of the mixing chamber interconnects the firstcompartment to the second compartment.
 14. A cartridge assemblyincluding the disposable cartridge of claim 1, further comprising: acapillary tube configured to introduce the sample fluid into thecartridge via the inlet; wherein at least one of the first reagentchamber and the mixing chamber is provided with an entry sealing member;wherein the entry sealing member prevents at least one of the firstreagent and fluid substance from flowing out of the at least one of thefirst reagent chamber and the mixing chamber via the inlet when theentry sealing member is intact; wherein the capillary tube is configuredto puncture the entry sealing member by inserting the capillary tubebeyond a position of the entry sealing member; and wherein the entrysealing member is configured to seal around a circumference of thecapillary tube when the capillary tube is inserted into the at least oneof the first reagent chamber and the mixing chamber.
 15. The cartridgeassembly of claim 14, wherein the mixing chamber comprises at least afirst compartment and a second compartment, wherein a narrow passage ofthe mixing chamber interconnects the first compartment to the secondcompartment.
 16. The cartridge assembly of claim 14, further comprising:the second reagent chamber being coupleable to an analyzing unit, whichperforms analysis of an output fluid; and a second frangible sealpreventing flow between the second reagent chamber and the analyzingunit, wherein the first and second frangible seals are configured toopen at different times.
 17. The cartridge assembly of claim 14, whereinat least one of the first reagent and the second reagent includes atleast one high molecular weight polymer in fluid form.
 18. The cartridgeassembly of claim 14, wherein the entry sealing member comprises: aninlet seal and a capillary seal, wherein the inlet seal prevents the atleast one of the first reagent and fluid substance from flowing out ofthe at least one of the first reagent chamber and the mixing chamber viathe inlet when the inlet seal is intact, the inlet seal configured to bepunctured by the capillary tube, and wherein the capillary seal isconfigured to seal around the circumference of the capillary tubeinserted through the capillary seal and into the at least one of thefirst reagent chamber and the mixing chamber.
 19. The cartridge assemblyof claim 18, wherein the inlet seal and capillary seal are provided insuccession within the inlet.
 20. The cartridge assembly of claim 14,wherein the capillary tube is coated with an anticoagulant.
 21. Thecartridge assembly of claim 20, wherein the anticoagulant isethylenediaminetetraacetic acid.
 22. The cartridge assembly of claim 14,further comprising: the capillary tube having a first end and a secondend; and a barrier positioned within the capillary tube, wherein thebarrier is configured to allow air inside of the capillary tube to exita second end of the capillary tube as the sample fluid flows into thecapillary tube through the first end and displaces the air inside of thecapillary tube, and wherein the barrier prevents passage of the samplefluid.
 23. The cartridge assembly of claim 22, wherein the barrier is ahydrophobic membrane.
 24. The cartridge assembly of claim 23, whereinthe hydrophobic membrane does not absorb the sample fluid.
 25. Thecartridge assembly of claim 22, wherein, after the sample fluid flowsinto the capillary tube through the first end, subsequent movement ofthe barrier towards the first end is configured to expel a predeterminedvolume of the sample fluid into the at least one of the first reagentchamber and the mixing chamber.
 26. A system including the cartridgeassembly of claim 25, further comprising: a plunger inserted into thesecond end of the capillary tube and advanced towards the first end ofthe capillary tube, thereby causing the subsequent movement of thebarrier towards the first end and the expelling of the sample fluid intothe at least one of the first reagent chamber and the mixing chamber.